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Metagenomic Analysis of Human Breast Milk
Breastfeeding is said to be the best source of infant nutrition. As a complete nutrition that is typically easiest for the baby to digest, it is highly recommended by most, if not all experts. It is said to promote healthy jaw strength and development, adequate digestion, and cognitive development as well as decrease the risk of allergies, diabetes, necrotizing enterocolitis, celiac disease, obesity, and SIDS in infants (1). Metagenomic Study Breast milk is deemed so beneficial because it includes bioactive components such as immune proteins and prebiotics which help to colonize a healthy gastrointestinal tract. This helps to combat adhesion and colonization of pathogenic bacteria while stimulating mucosal cell proliferation and enhancing immune development. This is all thanks to the wide array of bacteria found in the milk that benefit the body in a symbiotic manner. Scientists have performed a metagenomic study using 10 samples of different breast milk along with illumina sequencing and the MG-RAST pipeline in order to determine the different genera of bacteria, immune-modulatory DNA motifs, and the types of bacterial open reading frames in human milk that may influence bacterial establishment and stability of the GI tract (2). Results Metagenomic sequencing resulted in 261,532,204 51bp sequences read from the samples of breast milk. 1,331,996 of these sequences were of known prokaryotic genomes. More specifically, 75% were comprised of Staphylococcus, 15% of Pseudomonas, 2% of Edwardsiella, and 1% of Pantoea, Treponema, Streptococcus, and Campylobacter. The remaining 3% of known prokaryotic sequences mapped to 361 more bacterial genera, highlighting the immense diversity of the human milk metagenome. The sequences that were not identified as prokaryotic or human with very small basepair mismatches were realigned to the human genome with decreased stringency, resulting in 32,991,450 sequences left for contig assembly. They were then assembled and submitted to the MG-RAST pipeline, allowing a total of 53,285 sequences through quality control. Apon the analyzation of these sequences through MG-RAST, 65.1% were identified as Proteobacteria phyla while 34.6% were identified as Firmicute phyla. The contigs aligned to 194 known genomes, 61.1% being Pseudomonas, 33.4% being Staphylococcus, and 0.5% being ''Streptococcus ''(2). 33,793 open reading frames were analyzed and annotated using MG-RAST, while roughly 90% corresponded to a functional category. For example, many of the open reading frames encoded protein for basic cellular function such as respiration, cell signaling, RNA, DNA, and amino acid metabolism. Other open reading frames encoded proteins for stress response, carbohydrate metabolism, and virulence such as antibiotic resistance, adhesins, bacteriocins, and more. These functional open reading frames were also compared to fecal samples of the mothers, breast-fed infants, and formula-fed infants, all unrelated. Results found the greatest metagenomic variation between the human milk and the feces of formula-fed infants, highlighting 17 out of 26 functional categories that contained a significantly different proportion of open reading frames (2). These contigs were also searched for immunosuppressive motifs, showing the occurrence of sequences TTAAGGG and TCAAGCTTGA in human milk-contigs. The presence of these immunosuppressive motifs is similar to that in the metagenomes of breast-fed and formula-fed infants' feces as well as the mothers' feces. This suggests that possession of a diverse microbial community may lead to a similar abundance of immune suppressive motifs, regardless of the genera in the sample. Scientists also found that immunosuppressive motif TTAAGGG was in a higher abundance within the human genome than it was within bacteria. Because human breast milk contains macrophages, neutrophils, and natural killer cells, this suggests that ingestion of the maternal DNA by ingestion of immune cells and other free circulating DNA may contribute to proper immune development and regulation of infant immune response against a variety of bacterial, environmental, and food antigens. This is through exposure to immune stimulatory bacterial CpGs and immune suppressive DNA in both maternal and bacterial genomes (2). References 1. Wikipedia. "Breastfeeding". 2014. 2. Ward et al.: Human milk metagenome: a functional capacity analysis. BMC Microbiology 2013 13:116.